| RRC ID |
38852
|
| Author |
Kagami I, Mizunuma H, Miyamoto S, Ibuki Y, Uchida T.
|
| Title |
Quantification of estrogen receptor messenger RNA by quantitative polymerase chain reaction using internal standard fragment.
|
| Journal |
Biochem Biophys Res Commun
|
| Abstract |
A simple and reliable polymerase chain reaction-based method for quantifying human and rat estrogen receptor (ER)-mRNA is described. This method involves co-amplification of cDNA transcribed from sample RNA with an internal standard to reduce tube to tube amplification variations. Human and rat internal standards were synthesized by insertion of a DNA fragment near the midportion of human and rat target ER cDNA molecules. After co-amplification, both products of target ER cDNA and internal standard were separated on agarose gel by electrophoresis and transferred onto a membrane filter. The radioactivity hybridized with 32P-labeled ER cDNA was counted. The ER mRNA content was calculated by linear regression analysis after obtaining a logarithm of ratios between the sample and the internal standard radioactivity. The coefficient of variation of this assay was 18.8%. An absolute amount of ER mRNA in less than 10(4) cultured cells could be measured.
|
| Volume |
228(2)
|
| Pages |
358-64
|
| Published |
1996-11-12
|
| DOI |
10.1006/bbrc.1996.1666
|
| PII |
S0006-291X(96)91666-6
|
| PMID |
8920919
|
| MeSH |
Animals
Base Sequence
DNA Primers
DNA, Complementary
Endometrial Neoplasms / metabolism
Endometrium / metabolism
Estradiol / pharmacology
Female
Humans
Male
Mutagenesis, Insertional
Orchiectomy
Polymerase Chain Reaction / methods*
RNA, Messenger / analysis*
Rats
Rats, Wistar
Receptors, Estrogen / biosynthesis*
Recombinant Proteins / biosynthesis
Reference Standards
Reproducibility of Results
Transcription, Genetic / drug effects
Uterus / drug effects
Uterus / metabolism*
|
| IF |
2.985
|
| Times Cited |
5
|
|
WOS Category
|
BIOPHYSICS
BIOCHEMISTRY & MOLECULAR BIOLOGY
|
| Resource |
| Human and Animal Cells |
SKN(RCB0513)
HHUA(RCB0658)
HOS(RCB0992)
ROS-17/2.8-5(RCB0462) |
